Aimalohi Esechie

Protective effect of hydrogen sulfide in a murine model of acute lung injury induced by combined burn and smoke inhalation

Acute lung injury results in a severe inflammatory response, which leads to priming and activation of leucocytes, release of reactive oxygen and reactive nitrogen species, destruction of pulmonary endothelium, extravasation of protein-rich fluid into the interstitium and formation of oedema. Recently, H2S (hydrogen sulfide) has been shown to decrease the synthesis of pro-inflammatory cytokines, reduce leucocyte adherence to the endothelium and subsequent diapedesis of these cells from the microvasculature in in vivo studies, and to protect cells in culture from oxidative injury. In the present study, we hypothesized that a parenteral formulation of H2S would reduce the lung injury induced by burn and smoke inhalation in a novel murine model. H(2)S post-treatment significantly decreased mortality and increased median survival in mice. H2S also inhibited IL (interleukin)-1beta levels and significantly increased the concentration of the anti-inflammatory cytokine IL-10 in lung tissue. Additionally, H2S administration attenuated protein oxidation following injury and improved the histological condition of the lung. In conclusion, these results suggest that H2S exerts protective effects in acute lung injury, at least in part through the activation of anti-inflammatory and antioxidant pathways.

Beneficial effect of a hydrogen sulphide donor (sodium sulphide) in an ovine model of burn- and smoke-induced acute lung injury

Background and purpose:  The present study investigated whether the pathophysiological changes induced by burn and smoke inhalation are modulated by parenteral administration of Na2S, a H2S donor.

Assessment of vascular permeability in an ovine model of acute lung injury and pneumonia-induced Pseudomonas aeruginosa sepsis.

Objective:To assess the time changes and mechanism of pulmonary and peripheral vascular permeability in sheep with acute lung injury and sepsis. Design:Prospective, controlled, randomized trial. Setting:University research laboratory. Subjects:A total of 21 chronically instrumented, adult female sheep. Interventions:Sheep were instrumented with lung and prefemoral lymph fistulas and allocated to either an uninjured control group (n = 5) or sepsis group (n = 5). The sheep in the sepsis group received cotton smoke inhalation injury followed by instillation of Pseudomonas aeruginosa into the lungs. All sheep were mechanically ventilated and fluid resuscitated for the entire duration of the 24-hr experiment. Additional sheep (n = 11) received injury and were killed at different time points for the measurement of vascular endothelial growth factor in lung tissue. Measurements and Main Results:The injury induced a hypotensive–hyperdynamic circulation; increases in pulmonary capillary pressure, net fluid balance, lung and prefemoral lymph flow and protein content, lung water content, abdominal and thoracic fluid and protein content, neutrophil accumulation in the lung, and vascular endothelial growth factor expression in lung tissue; and decreases in Pao2/Fio2 ratio, plasma protein concentration, plasma oncotic pressure, and myocardial contractility. Conclusions:Lung edema formation in this model was the result of marked increases in both pulmonary microvascular permeability and pressure. Pulmonary vascular hyperpermeability peaked 12 hrs postinjury and was related to vascular endothelial growth factor overexpression. Early myocardial failure was a potential contributor to the constant increase in pulmonary capillary pressure. The sepsis-induced increase in peripheral microvascular permeability was associated with significant accumulation of fluid and protein in the third space.

Combined anticoagulants ameliorate acute lung injury in sheep after burn and smoke inhalation.

Burn and smoke inhalation-related multiple organ dysfunction is associated with a severe fall in the plasma concentration of antithrombin. Therefore the aim of the present study was to test the hypothesis that intravenous administration of recombinant human antithrombin in combination with aerosolized heparin will ameliorate acute lung injury in sheep exposed to cutaneous burn and smoke inhalation. Sheep were prepared operatively for study and, 7 days post-surgery, sheep were given a cutaneous burn (40% of total body surface area, third-degree burn) and insufflated with cotton smoke (48 breaths, <40 degrees C) under halothane anaesthesia. After injury, sheep were placed on a ventilator and resuscitated with Ringers lactate solution. The animals were divided into three groups: sham group (non-injured and non-treated; n=6), saline group (injured and received saline; n=6) and rhAT.iv.+Hep group [injured and treated with rhAT (recombinant human antithrombin) and heparin; n=6]. In the rhAT.iv.+Hep group, rhAT was infused continuously for 48 h starting 1 h post-injury with a dose of 0.34 mg.h(-1).kg(-1) of body weight and heparin (10000 units) was aerosolized every 4 h starting at 1 h post-injury. The experiment lasted 48 h. Haemodynamics were stable in sham group, whereas the saline-treated sheep developed multiple signs of acute lung injury, including decreased pulmonary gas exchange, increased inspiratory pressures, extensive airway obstruction and increased pulmonary oedema. These pathological changes were associated with a severe fall in plasma antithrombin concentration, lung tissue accumulation of leucocytes and excessive production of NO. Treatment of injured sheep with anticoagulants attenuated all of the pulmonary pathophysiology observed. In conclusion, the results provide definitive evidence that anticoagulant therapy may be a novel and effective treatment tool in the management of burn patients with concomitant smoke inhalation injury.

Time course of nitric oxide synthases, nitrosative stress, and poly(ADP ribosylation) in an ovine sepsis model

IntroductionDifferent isoforms of nitric oxide synthases (NOS) and determinants of oxidative/nitrosative stress play important roles in the pathophysiology of pulmonary dysfunction induced by acute lung injury (ALI) and sepsis. However, the time changes of these pathogenic factors are largely undetermined.MethodsTwenty-four chronically instrumented sheep were subjected to inhalation of 48 breaths of cotton smoke and instillation of live Pseudomonas aeruginosa into both lungs and were euthanized at 4, 8, 12, 18, and 24 hours post-injury. Additional sheep received sham injury and were euthanized after 24 hrs (control). All animals were mechanically ventilated and fluid resuscitated. Lung tissue was obtained at the respective time points for the measurement of neuronal, endothelial, and inducible NOS (nNOS, eNOS, iNOS) mRNA and their protein expression, calcium-dependent and -independent NOS activity, 3-nitrotyrosine (3-NT), and poly(ADP-ribose) (PAR) protein expression.ResultsThe injury induced severe pulmonary dysfunction as indicated by a progressive decline in oxygenation index and concomitant increase in pulmonary shunt fraction. These changes were associated with an early and transient increase in eNOS and an early and profound increase in iNOS expression, while expression of nNOS remained unchanged. Both 3-NT, a marker of protein nitration, and PAR, an indicator of DNA damage, increased early but only transiently.ConclusionsIdentification of the time course of the described pathogenetic factors provides important additional information on the pulmonary response to ALI and sepsis in the ovine model. This information may be crucial for future studies, especially when considering the timing of novel treatment strategies including selective inhibition of NOS isoforms, modulation of peroxynitrite, and PARP.

Combined neuronal and inducible nitric oxide synthase inhibition in ovine acute lung injury.

Objective:Acute lung injury with subsequent pneumonia and sepsis represents a major cause of morbidity and mortality in thermally injured patients. Production of nitric oxide by the neuronal and inducible nitric oxide synthase may be critically involved in the pathophysiology of the disease process at different time points, and thus specific inhibition at different times may represent an effective treatment regimen. Design:Prospective, controlled, randomized trial. Setting:University research laboratory. Subjects:Eighteen chronically instrumented, adult, female sheep. Interventions:Following baseline measurements, the animals were allocated to either sham-injured, nontreated controls (sham), injured, nontreated controls (control), or injured animals treated with continuous infusion of 7-nitroindazole, a specific neuronal nitric oxide synthase inhibitor, during the first 12 hrs postinjury and infusion of BBS-2, a specific inducible nitric oxide synthase inhibitor, during the next 12 hrs. Injury was induced by 48 breaths of cotton smoke and subsequent instillation of Pseudomonas aeruginosa into the lungs. All sheep were mechanically ventilated and fluid resuscitated for the entire duration of the 24-hr experiment. Measurements and Main Results:The injury induced severe pulmonary dysfunction, which was associated with increases in lung edema formation, airway obstruction, and vascular endothelial growth factor, 3-nitrotyrosine, and poly(adenosine diphosphate ribose) expression in lung tissue. The treatment reduced the degree of airway obstruction and improved pulmonary gas exchange, whereas the development of lung edema was not affected. The increases in lung tissue vascular endothelial growth factor, 3-nitrotyrosine, and poly(ribose) expression were attenuated by the treatment. Conclusions:The combination of early neuronal nitric oxide synthase and delayed inducible nitric oxide synthase inhibition shows potential benefit in ovine acute lung injury by reducing nitrosative stress in the lung and limiting the degree of airway obstruction.

Role of different nitric oxide synthase isoforms in a murine model of acute lung injury and sepsis

Excessive production of nitric oxide (NO) by NO synthase (NOS) with subsequent formation of peroxynitrite and poly(adenosine diphosphate ribose) is critically implemented in the pathophysiology of acute lung injury and sepsis. To elucidate the roles of different isoforms of NOS, we tested the effects of non-selective NOS inhibition and neuronal NOS (nNOS)- and inducible NOS (iNOS)-gene deficiency on the pulmonary oxidative and nitrosative stress reaction in a murine sepsis model. The injury was induced by four sets of cotton smoke using an inhalation chamber and subsequent intranasal administration of live Pseudomonas aeruginosa (3.2x10(7) colony-forming units). In wild type mice, the injury was associated with excessive releases of pro-inflammatory cytokines in the plasma, enhanced neutrophil accumulation, increased lipid peroxidation, and excessive formation of reactive nitrogen species and vascular endothelial growth factor in the lung. Both nNOS- and iNOS-gene deficiency led to significantly reduced oxidative and nitrosative stress markers in the lung, but failed to significantly improve survival. Treatment with a non-selective NOS inhibitor failed to reduce the oxidative and nitrosative stress reaction to the same extent and even tended to increase mortality. In conclusion, the current study demonstrates that both nNOS and iNOS are partially responsible for the pulmonary oxidative and nitrosative stress reaction in this model. Future studies should investigate the effects of specific pharmacological inhibition of nNOS and iNOS at different time points during the disease process.

A murine model of sepsis following smoke inhalation injury

Acute lung injury (ALI) by smoke inhalation with subsequent pneumonia and sepsis represents a major cause of morbidity and mortality in burn patients. The aim of the present study was to develop a murine model of ALI and sepsis to enhance the knowledge of mechanistic aspects and pathophysiological changes in patients with these injuries. In deeply anesthetized female C57BL/6 mice, injury was induced by four sets of cotton smoke using an inhalation chamber. Afterward, live Pseudomonas aeruginosa (3.2x10(7) colony-forming units) was administered intranasally. The indicated dose of bacteria was determined based on the results of a dose-response study (n=47). The following study groups were monitored for survival over 96h: (1) sham injury group, (2) only smoke inhalation group, (3) only bacteria group, and (4) smoke inhalation plus bacteria group. Each group included 10 mice. The survival rates were 100%, 90%, 30%, and 10%, respectively. The double hit injury was associated with excessive releases of pro-inflammatory cytokines in the plasma, and enhanced neutrophil accumulation, increased lipid peroxidation, and excessive formation of reactive nitrogen species in the lung. In mice receiving only smoke inhalation injury, no systemic cytokine release and increased lung tissue lipid peroxidation were observed. However, smoke alone significantly increased neutrophil accumulation and formation of reactive nitrogen species in lung tissue. In conclusion, bacterial pneumonia is predominantly responsible for mortality and morbidity in this novel murine model of smoke inhalation and pulmonary sepsis. Reactive oxygen and nitrogen species mediate the severity of lung injury.

Effects of early neuronal and delayed inducible nitric oxide synthase blockade on cardiovascular, renal, and hepatic function in ovine sepsis.

Background:Recent evidence suggests that nitric oxide produced via the neuronal nitric oxide synthase is involved mainly in the early response to sepsis, whereas nitric oxide derived from the inducible nitric oxide synthase is responsible during the later phase. We hypothesized that early neuronal and delayed inducible nitric oxide synthase blockade attenuates multiple organ dysfunctions during sepsis. Methods:Sheep were randomly allocated to sham-injured, nontreated animals (n = 6); injured (48 breaths of cotton smoke and instillation of Pseudomonas aeruginosa into the lungs), nontreated animals (n = 7); and injured animals treated with a neuronal nitric oxide synthase inhibitor from 1 to 12 h and an inducible nitric oxide synthase inhibitor from 12 to 24 h postinjury (n = 6). Results:The injury induced arterial hypotension, vascular leakage, myocardial depression, and signs of renal and hepatic dysfunctions. The treatment significantly attenuated, but did not fully prevent, the decreases in mean arterial pressure and left ventricular stroke work index. Although the elevation of creatinine levels was partially prevented, the decreases in urine output and creatinine clearance were not affected. The injury-related increases in bilirubin levels, international normalized ratio, and lipid peroxidation in liver tissue were significantly attenuated. Although plasma nitrite/nitrate levels were significantly increased versus baseline from 12–24 h in controls, plasma nitrite/nitrate levels were not increased in treated animals. Conclusions:The combination treatment shows potential benefit on sepsis-related arterial hypotension and surrogate parameters of organ dysfunctions in sheep. It may be crucial to identify the time course of expression and activation of different nitric oxide synthase isoforms in future investigations.

Cardiovascular collapse and vascular permeability changes in an ovine model of methicillin-resistant Staphylococcus aureus sepsis.

Methicillin-resistant Staphylococcus aureus (MRSA) infections with severe outcomes such as sepsis and septic shock are progressively increasing in both the community and in hospital settings. We hypothesized that overexpression of reactive nitrogen and oxygen species and vascular endothelial growth factor (VEGF) play a pivotal role in cardiovascular collapse associated with vascular hyperpermeability in MRSA sepsis. Twelve sheep were surgically prepared and randomized into a control (noninjured; n = 6) and a sepsis (injured; n = 6) group. Animals in the sepsis group were subjected to cotton smoke inhalation and instillation of 2.5 × 1011 colony-forming units of live MRSA into both lungs. Cardiovascular variables in the control group remained stable, whereas the MRSA sepsis group developed a hypotensive and hyperdynamic circulatory shock state beginning at 6 h associated with significantly increased vascular permeability evidenced by increased prefemoral lymph flow starting at 12 h and permeability index from 12 to 18 h, higher fluid accumulation from 12 to 24 h, and significantly decreased plasma protein concentration and oncotic pressure beginning at 6 h compared with control animals. Myocardial 3-nitrotyrosine (3-NT) protein, poly (adenosine diphosphate-ribose), and VEGF mRNA expressions measured after the 24-h experiment were significantly increased in the injured animals as well. These results evidence that excessive production of reactive radicals and VEGF may play a major role in cardiovascular collapse and vascular hyperpermeability in MRSA sepsis.